Proposal for Standardization of Optimized Mycobacterial Interspersed Repetitive Unit-Variable-Number Tandem Repeat Typing of Mycobacterium tuberculosis

Author:

Supply Philip12,Allix Caroline3,Lesjean Sarah12,Cardoso-Oelemann Mara12,Rüsch-Gerdes Sabine4,Willery Eve12,Savine Evgueni12,de Haas Petra5,van Deutekom Henk6,Roring Solvig7,Bifani Pablo1238,Kurepina Natalia8,Kreiswirth Barry8,Sola Christophe9,Rastogi Nalin9,Vatin Vincent210,Gutierrez Maria Cristina11,Fauville Maryse3,Niemann Stefan4,Skuce Robin7,Kremer Kristin4,Locht Camille12,van Soolingen Dick4

Affiliation:

1. INSERM, U629

2. Institut Pasteur de Lille, Lille, France

3. Institut Pasteur de Bruxelles, Laboratoire Tuberculose et Mycobactéries, Brussels, Belgium

4. Forschungszentrum Borstel, National Reference Center for Mycobacteria, Borstel, Germany

5. Diagnostic Laboratory for Infectious Diseases and Perinatal Screening, National Institute for Public Health and the Environment, 3720 Bilthoven

6. Department of Tuberculosis Control, Municipal Health Service, Amsterdam, The Netherlands

7. Veterinary Sciences Division, Department of Agriculture and Rural Development, Belfast, Northern Ireland

8. TB Center, Public Health Research Institute, Newark, New Jersey

9. Unité de la Tuberculose et des Mycobactéries, Institut Pasteur de Guadeloupe, Guadeloupe

10. CNRS, UMR 8090, Lille, France

11. Laboratoire de Référence des Mycobactéries, Institut Pasteur, Paris, France

Abstract

ABSTRACT Molecular typing based on 12 loci containing variable numbers of tandem repeats of mycobacterial interspersed repetitive units (MIRU-VNTRs) has been adopted in combination with spoligotyping as the basis for large-scale, high-throughput genotyping of Mycobacterium tuberculosis . However, even the combination of these two methods is still less discriminatory than IS 6110 fingerprinting. Here, we define an optimized set of MIRU-VNTR loci with a significantly higher discriminatory power. The resolution and the stability/robustness of 29 loci were analyzed, using a total of 824 tubercle bacillus isolates, including representatives of the main lineages identified worldwide so far. Five loci were excluded for lack of robustness and/or stability in serial isolates or isolates from epidemiologically linked patients. The use of the 24 remaining loci increased the number of types by 40%—and by 23% in combination with spoligotyping—among isolates from cosmopolitan origins, compared to those obtained with the original set of 12 loci. Consequently, the clustering rate was decreased by fourfold—by threefold in combination with spoligotyping—under the same conditions. A discriminatory subset of 15 loci with the highest evolutionary rates was then defined that concentrated 96% of the total resolution obtained with the full 24-locus set. Its predictive value for evaluating M. tuberculosis transmission was found to be equal to that of IS 6110 restriction fragment length polymorphism typing, as shown in a companion population-based study. This 15-locus system is therefore proposed as the new standard for routine epidemiological discrimination of M. tuberculosis isolates and the 24-locus system as a high-resolution tool for phylogenetic studies.

Publisher

American Society for Microbiology

Subject

Microbiology (medical)

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